TRAF4 overexpression is a common characteristic of human carcinomas.

Tumor necrosis factor receptor (TNFR) associated factor 4 (TRAF4) was initially identified as a gene amplified and overexpressed in breast carcinomas. Our aim was to evaluate whether TRAF4 protein overexpression exists in other cancer types. Immunohistochemistry analysis of tumor samples from 623 patients with 20 different tumor types showed that TRAF4 was overexpressed in 268 tumors (43%), including 82 of 137 lung adenocarcinomas (60%). Interestingly, 32 primary tumors and their matching metastases exhibited mostly similar TRAF4 expression pattern. TRAF4 protein overexpression was limited to cancer cells and the subcellular localization was consistently cytoplasmic in a large majority of cases. To investigate changes in TRAF4 gene copy number, 125 cases from six different types of carcinomas were also analysed by fluorescence in situ hybridization. Out of the 28 cases (22%) showing an increased TRAF4 gene copy number, 23 (82%) were overexpressing the protein. Thus, TRAF4 gene amplification is one of the mechanisms responsible for TRAF4 protein overexpression in human cancers. Considering that TRAF4 is located at 17q11.2 in a region of amplification devoid of known oncogenes and is commonly overexpressed in cancer, our data support an oncogenic role for TRAF4.

Common 4q24 deletion in four cases of hematopoietic malignancy: early stem cell involvement?

We determined bone marrow karyotype at diagnosis in four female acute myeloid leukemia (AML) or myelodysplasia patients, aged between 52 and 56 years. In each case, we observed chromosome rearrangement involving the same 4q24 band. Three patients had a balanced reciprocal translocation as the sole abnormality - t(3;4)(q26;q24), t(4;5)(q24;p16) and t(4;7)(q24;q21) - and the fourth had del(4)(q23q24), +4. We used a set of 4q BAC probes for fluorescent in situ hybridization (FISH) in these four cases. We found a 4q24 submicroscopic deletion in all three translocations, with a common deletion of approximately 0.5 Mb. In three cases, we concluded that rearrangement occurred in an early hematopoietic stem cell, as it was detected, in mosaic with a normal karyotype, in a fraction of remission bone marrow cells, peripheral T and B lymphocytes, malignant lymph node T-lymphoma cells in one case and B-lymphoblastoid cell lines established in two cases. Moreover, one of 10 additional AML patients tested by FISH had a normal karyotype and deletion of one of the commonly deleted probe sequences. A tumor suppressor gene may therefore be involved, especially as two patients developed malignant lymphoma at the same time as myeloid proliferation.

Multivariate analysis of prognostic factors in acute myeloid leukemia: value of clonogenic leukemic cell properties.

The initial clinical and biological parameters, including clonogenic leukemic cell (CFU-L) assay, were reviewed for their prognostic significance in a cohort of 188 adult patients with newly diagnosed untreated acute myeloid leukemia (AML). Almost all patients received induction therapy with daunorubicin (DNR) and cytarabine (Ara-C) according to the European Organization for Research and Treatment of Cancer (EORTC) AML 5 to AML 9 trials. Bone marrow samples from 116 representative patients were obtained for CFU-L assay with an efficiency percentage of 89.6%; 76 patients had a measurement of the CFU-L self-renewal capacity (second plating efficiency [PE2]) and 91 patients had CFU-L inhibition test after exposure to DNR and/or Ara-C. The prognostic significance of parameters such as age, hematological antecedent, WBC count, liver enlargement, and Auer rods is confirmed in the present study. Moreover, high platelet and polymorphonuclear counts appeared to be related to resistance to induction course. However, through multivariate analysis, CFU-L sensitivity to drugs and self-renewal capacity appeared to be major independent prognostic factors in AML. A low CFU-L inhibition in the presence of the DNR and Ara-C combination correlates with a poorer complete remission (CR) rate, but not with CR duration. Patients with the lower PE2 values experienced both higher CR rate and longer CR duration. The practical interest of CFU-L study remains to be defined but, at least, PE2 measurement could be considered in the future as a major variable in determining therapeutic aggressiveness.

Fluorescence in situ hybridization analysis of minute marker chromosomes in leukemia with monosomy 7.

Monosomy 7 was detected in bone marrow cells from three patients, one with myeloid leukemia, and two others with myelodysplastic syndrome following previous chemotherapy. Fluorescence in situ hybridization (FISH), carried out with an alphoid DNA probe specific for chromosome 7 centromere, showed that a small marker chromosome present in the tumor cells' karyotype of the three patients, was derived from the missing chromosome 7. In two cases, the marker was a ring chromosome, whereas in the third case it was a tiny dot-like chromosome, unnoticed at first examination on R-banded metaphases. In the three cases, the marker was lost in a proportion of tumor cells. FISH experiments suggested that the marker centromere had undergone structural alterations, with a fluorescence pattern distinct from a normal one. On the whole, these data suggest that: firstly, leukemia-associated monosomy 7 results, in a proportion of cases, from a structural event rather than from simple loss of a whole chromosome 7; secondly, interpretation of interphase FISH must be cautious in monosomy 7 evaluation; and thirdly structural alteration of the chromosome 7 derivative alphoid DNA could explain its propensity to segregate unequally and to be lost at mitosis.

Thrombocytemia and abnormal megakaryopoiesis associated with abnormality of chromosome 1p34 in myelodysplastic syndromes.

Myelodysplastic syndromes (MDS) are a heterogeneous group of diseases characterized by a cytopenia related to abnormal proliferation and differentiation of marrow precursor cells. Some subtypes of MDS are associated with thrombocytemia or abnormal megakaryopoiesis and certain specific karyotypic abnormalities. We report on four cases of MDS with normal or elevated blood platelet count and a recurring abnormality in chromosome 1p34. The gene involved appears to be different from c-mpl and TPO.

Prognostic value of c-mpl expression in myelodysplastic syndromes.

The c-mpl proto-oncogene which encodes a member of the hematopoietic cytokine receptor superfamily has been recently shown to be the receptor for thrombopoietin (TPO), which stimulates megakaryocyte progenitor expansion and differentiation. We studied c-mpl expression by Northern blot analysis, in a large series of 58 MDS. No expression was found in 14 patients with refractory anemia (RA) or with refractory anemia with ring sideroblasts (RARS). In contrast 11/26 (42%) patients with refractory anemia with excess of blasts (RAEB), or with RAEB in transformation (RAEBt), and 8/18 (44%) patients with chronic myelomonocytic leukemia (CMML) expressed c-mpl. In CMML patients, no correlation was found between c-mpl expression and any prognostic factor tested, nor with the course of the disease. In contrast, in RAEB and RAEBt, expression of c-mpl was correlated with high Bournemouth scoring (P < 0.005) and poor survival (P = 0.02) due to leukemic transformation. Forty-five per cent (5/11) of the c-mpl positive patients evolved towards AML with a mean follow-up of 10.5 months, while 13% (2/15) of the c-mpl negative patients developed a secondary leukemia, with a mean follow-up of 21.1 months. Moreover, in RAEB and RAEBt, a significant correlation was observed between c-mpl, CD34, megakaryocyte glycoprotein IIb (GPIIb) expression, and the presence of dysmegakaryopoiesis. These results indicate that patients with RAEB and RAEBt, with high expression of the c-mpl, CD34, and GPIIb genes, may identify a subgroup of patients with particularly poor prognosis, due to an increased risk of secondary leukemia. More aggressive therapy could be justified in these patients.

Report of 34 patients with clonal chromosomal abnormalities in Philadelphia-negative cells during imatinib treatment of Philadelphia-positive chronic myeloid leukemia.

Imatinib mesylate (Gleevec), an inhibitor of the BCR-ABL tyrosine kinase, was introduced recently into the therapy of chronic myeloid leukemia (CML). Several cases of emergence of clonal chromosomal abnormalities after therapy with imatinib have been reported, but their incidence, etiology and prognosis remain to be clarified. We report here a large series of 34 CML patients treated with imatinib who developed Philadelphia (Ph)-negative clones. Among 1001 patients with Ph-positive CML treated with imatinib, 34 (3.4%) developed clonal chromosomal abnormalities in Ph-negative cells. Three patients were treated with imatinib up-front. The most common cytogenetic abnormalities were trisomy 8 and monosomy 7 in twelve and seven patients, respectively. In 15 patients, fluorescent in situ hybridization with specific probes was performed in materials archived before the initiation of imatinib. The Ph-negative clone was related to previous therapy in three patients, and represented a minor pre-existing clone that expanded after the eradication of Ph-positive cells with imatinib in two others. However, in 11 patients, the new clonal chromosomal abnormalities were not detected and imatinib may have had a direct effect. No myelodysplasia was found in our cohort. With a median follow-up of 24 months, one patient showed CML acceleration and two relapsed.

Occupational risk factors for acute leukaemia: a case-control study.

A case-control study has been performed for occupational risk factors of acute leukaemia, based on 185 cases more than 30 years old and 513 matched controls. There was a significant excess of polyvalent farming and electronic engineers among professions of cases, and, in addition of metal workers when considering the professions pursued for more than 5 years. The corresponding exposures were analysed through a detailed questionnaire, and assessed by an industrial hygienist after blinding the case-control status. The odds ratios (OR) were computed after adjustment on matching variables and prior chemo- or radiotherapy treatment, and after stratification for the level and total duration of exposure. There was no excess of professional exposure to ionizing radiation among cases. A significant relationship was observed between acute leukaemia and high or medium exposure to benzene, as well as over 10 years high or medium exposure to exhaust gas. In addition a significant relationship was observed with exposure to pesticides--insecticides and/or weed killers--and to electric and magnetic fields (EMF). The relationship with pesticides was significant when considering high or medium exposure to weed killers and more than 10 years exposure to both subtypes of pesticides. The relationship with pesticides and EMF remained significant when confounding factors were taken into consideration and after adjustment on co-exposure to benzene. The cytological studies showed that acute leukaemias following exposure to benzene (high or medium) and to EMF were only of myelogenous subtypes, whereas those following exposure to pesticides were divided between lymphoblastic and myeloblastic subtypes. Cytogenetic studies failed to show increased frequency of chromosomal abnormalities, as described in acute leukaemias secondary to anti-cancer treatments. Our study adds credence to the hypothesis that pesticides and EMF are leukaemogenic agents, together with benzene.

Three cases of preleukemic myelodysplastic disorders with the same translocation t(1;3).

The same translocation, t(1;3)(p36;q21), was observed in three patients with a preleukemic syndrome presenting as a myelodysplastic disorder. It was the only chromosomal abnormality in two patients; and the third patient had 12p- and 15q- as additional abnormalities. The three cases evolved into acute nonlymphocytic leukemia.

Acute myelomonocytic leukemia in a XYY man.

A case of acute myeloid leukemia (M4) in a 29-year-old male with a 47,XYY karyotype is reported. This aneuploidy was found in both bone marrow cells and mitogen-stimulated lymphocytes. Monosomy 7 correlated with myelodysplastic features. The possible role of XYY in increasing the risk of leukemia is discussed.

Another case of trisomy 4 with double minute chromosomes in acute non-lymphocytic leukemia.

A further case of trisomy 4 with double minute chromosomes in acute non-lymphocytic leukemia is reported. The non-random association between these two cytogenetic abnormalities is reinforced. A possible relation with environmental exposure is discussed.

Submicroscopic insertion of RARalpha gene into chromosome 15 in two cases of acute promyelocytic leukemia.

Acute promyelocytic leukemia (APL) is characterized by a specific translocation (15;17)(q22;q21), resulting in the formation of PML/RARalpha chimeric transcripts. We report two female patients with PML/RARalpha-positive classical APL, whose leukemic cells expressed a variant translocation, t(5;15)(q13;q22) and t(15;17)(q22;p13), respectively. Both translocations were confirmed by whole chromosome painting which revealed no apparent involvement of 17q. A two-color fluorescence in situ hybridization with a 5' PML and a 3' RARalpha probe showed, in both cases, the presence of a PML-RARalpha fusion gene, on the der(15)t(5;15) long arm, and on the der(17)t(15;17) short arm, respectively. These two complex rearrangements resulted most probably from a two-step mechanism: (1) a submicroscopic insertion into 15q of a 17q segment including the 3' part of the RARalpha gene; (2) a reciprocal translocation between der(15) and a variable chromosome arm, with a breakpoint distal and proximal to RARalpha insertion in the case of t(5;15) and t(15;17), respectively. Molecular and prognosis significance of these variant translocations are discussed.

Pentasomy 13q in a case of acute myelogenous leukemia (Mo).

A case of acute myelogenous leukemia Mo FAB subtype with a pentasomy 13q (associated with a trisomy 19 in a subclone) in the initial bone marrow metaphase cells is reported. The pentasomy 13q is the result of the presence of double isochromosome 13q and one normal chromosome 13. In our case, this abnormality had a poor prognosis.

Aplastic anemia responsive to cyclosporine complicating the evolution of polycythemia vera.

We report here a very unusual patient with Polycythemia vera treated with Pipobroman who developed severe aplastic anemia following administration of the drug. Six months later, because of lack of response, cyclosporine therapy was given there was rapid and complete hematological recovery, highly suggestive of an immune-mediated mechanism, in this case.

Control of cancer-related pain with MS Contin: a comparison between 12-hourly and 8-hourly administration.

Nineteen cancer patients with chronic pain of moderate to severe intensity were randomized in a double-blind manner to 5 days of either 8-hourly or 12-hourly administration of controlled-release morphine (MS Contin, MSC), followed by the alternate schedule for 5 days. The control of pain, using an average dose of 303.4 +/- 254.4 mg/day of MSC, was good during both the 8-hourly and 12-hourly phases, and the mean daily pain intensity measured by visual analogue scale (VAS), pain relief (VAS), and global efficacy scores did not differ when compared by treatment schedule. The need for supplemental "rescue" morphine was infrequent and did not differ between treatment phases (8-hourly, 0.7 +/- 0.7 and 12-hourly, 0.6 +/- 0.6 doses per day, p = 0.6232). The overall frequency and severity of adverse events did not differ between the two dosing schedules. A majority of patients (67%) reported that they believed that 12-hourly dosing was a moderate or great advantage over 8-hourly dosing.

PI-103, a dual inhibitor of Class IA phosphatidylinositide 3-kinase and mTOR, has antileukemic activity in AML.

The phosphatidylinositol 3-kinase (PI3K)/Akt and mammalian target of rapamycin complex 1 (mTORC1) signaling pathways are frequently activated in acute myelogenous leukemia (AML). mTORC1 inhibition with RAD001 induces PI3K/Akt activation and both pathways are activated independently, providing a rationale for dual inhibition of both pathways. PI-103 is a new potent PI3K/Akt and mTOR inhibitor. In human leukemic cell lines and in primary blast cells from AML patients, PI-103 inhibited constitutive and growth factor-induced PI3K/Akt and mTORC1 activation. PI-103 was essentially cytostatic for cell lines and induced cell cycle arrest in the G1 phase. In blast cells, PI-103 inhibited leukemic proliferation, the clonogenicity of leukemic progenitors and induced mitochondrial apoptosis, especially in the compartment containing leukemic stem cells. In contrast, apoptosis was not induced with RAD001 and IC87114 association, which specifically inhibits mTORC1 and p110delta activity, respectively. PI-103 had additive proapoptotic effects with etoposide in blast cells and in immature leukemic cells. Interestingly, PI-103 did not induce apoptosis in normal CD34(+) cells and had moderate effects on their clonogenic and proliferative properties. Here, we demonstrate that multitargeted therapy against PI3K/Akt and mTOR with PI-103 may be of therapeutic value in AML.

[Use of the concanavalin-peroxidase-DAB system for identification of human chromosomes]. / Utilisation du système concanavaline-peroxydase-DAB pour l'identification des chromosomes humains.

The effect of concanavalin A on the prometaphase chromosomes was investigated, using a staining reaction based on the peroxydase-diaminobenzidin-H2O2 system. After incubation with concanavalina A, the chromosomes telomeres as well as the centromeres and satellites of the acrocentric chromosomes were strongly stained. Sometimes the chromatids appeared to be coiled. In other respect, it must be noted that peroxydase alone can stain the chromatids, which probably means that this compounds is able to unite with the chromosomes, without the aid of concanavalin A.